PROJECT SUMMARY/ABSTRACT Adenoviruses (AdV) are common human pathogens that cause typical cold symptoms in healthy indi- viduals, but can potentially progress to acute respiratory distress syndrome (ARDS) with up to 50% mortality, particularly in highly susceptible, immunosuppressed people, and new, potentially lethal variants continue to emerge each year. No therapeutic that specifically prevents or treats AdV infection exists and the development of novel treatments would prevent the morbidity and potentially mortality associated with AdV infection. The objective of this proposal is to determine the mechanism of action and anti-adenovirus efficacy of novel molecules that decrease apical Coxsackievirus and adenovirus receptor (CAREx8) protein expression. Our central hypothesis is that decoy peptides that block the interaction between MAGI-1 and CAREx8 destabi- lize apical CAREx8 protein in the airway to abrogate AdV entry and pathogenesis. We will test our hypothesis using polarized model epithelia with Dox-inducible CAREx8 expression, well-differentiated primary human and rodent airway epithelia to validate our findings, and the cotton rat model to evaluate wildtype AdV pathogenesis in immunocompetent and cyclophosphamide-immunosuppressed animals, with two specific aims: Aim 1: To elucidate the mechanism of MAGI-1 activating peptide and AdV triggered proteolytic degra- dation of CAREx8. Completion of this aim will allow the identification and development of novel and much needed targets and approaches to prevent the infection and spread of pathogenic wild-type AdV and, in the future, group B coxsackieviruses. Aim 2: To define the protection afforded by MAGI-1 PDZ1 binding peptides against wild type adenovi- rus infection in cotton rats. Completion of this aim will provide proof-of-principle that AdV infection can be thwarted by reducing its primary receptor and may save lives of severely infected or immunosuppressed indi- viduals. Overall impact: Understanding the mechanisms that regulate the expression and localization of CAREx8, and how this unique isoform mediates viral entry at the apical surface of a polarized epithelium is criti- cal for understanding viral spread, tissue tropism, and pathogenesis. The successful completion of the pro- posed aims will identify not only the cellular mechanisms regulating the expression of the apical adenovirus receptor but also mechanisms regulating viral binding and infection. This will establish the feasibility of thera- peutic agents that reduce the susceptibility of the airway epithelium to adenovirus infection and pathogenicity prior to infection and during an active infection in immunocompetent and immunosuppressed conditions. We ultimately expect the proposed aims to lead to novel anti-viral interventions that may also block other viruses that use CAR as a primary receptor, and provide insight into the regulation of other related viral receptors.